Early detection of cyanotoxin threat in freshwater bodies by quantitative PCR

Increased frequency of cyanobacterial blooms, caused by rising temperatures and eutrophication, is creating a growing need for reliable methods for early cyanotoxin threat detection in water bodies. Current methods are time-consuming and sensitive to taxonomic inconsistencies. On top of that, they only enable detection of potentially toxin-producing cells when they have already multiplied to a relatively large number. To develop a tool for earlier detection of potentially toxic cyanobacterial strains, we are using qPCR to target genes involved in crucial parts of cyanotoxin synthesis. Detection and quantification of these genes, if incorporated in regular monitoring, might serve as an early warning in case of increased potential for cyanotoxin production. Toxins and genes of interest in this study are microcystins (mcyE), saxitoxins (sxtA) and cylindrospermopsins (cyrJ). The analyses will be carried out on 28 phytoplankton and 23 phytobenthos samples from 15 different freshwater bodies in Slovenia. This way we will include also understudied benthic cyanobacteria and their ability to produce toxic metabolites. Results will be compared with taxa list obtained by traditional microscopic identification methods and with cyanotoxin content evaluated by LC-MS/MS. The study has a potential to prepare the ground for improving current monitoring programs by complementing them with highly sensitive molecular methods. This can provide more in-depth information about dynamics of toxic cyanobacterial populations in water bodies and thus help with adopting appropriate mitigation strategies

Occurrence of cyanobacterial toxins in freshwaters of the Alpine region

Different genera of freshwater cyanobacteria have the ability of producing toxic metabolites (cyanotoxins). Massive proliferation of these organisms can lead to harmful concentration of toxins in the water. An increase in frequency, magnitude and duration of harmful blooms has been documented globally in the last decades. It is therefore important to gather information about the occurrence of these contaminants in order to understand their geographical spread. The distribution of cyanotoxins in freshwater bodies of the alpine region was investigated in the frame of the “EcoAlps-Water '' project, funded by the European Interreg Alpine Space Program. The sampling was conducted in lakes and rivers of Austria, Italy, Slovenia and Switzerland using uniform protocols. Both pelagic and benthic samples were sampled. Toxins’ analysis was conducted in a reference laboratory (at the Fondazione Edmund Mach) using LC-MS/MS methodologies, able to distinguish the different classes of toxic compounds: hepatotoxic microcystins, neurotoxic anatoxins and saxitoxins, cytotoxic cylindrospermopsins. The investigation revealed the presence of cyanotoxins in the majority of investigated lakes, with microcystins being the most represented. Moreover, anatoxins were found in a few water bodies. In conclusion, cyanotoxins represent an additional risk that lakes of the Alpine Space region are facing. Constant attention to this issue must therefore be paid because global climatic changes and other local anthropogenic pressures can synergistically increase the risk of high cyanotoxins levels

Chemical synthesis and biological activities of 3-alkyl pyridinium polymeric analogues of marine toxins

Two new large poly-1,3-dodecylpyridinium salts, APS12 and APS12-2 of 12.5- and 14.7-kDa size, respectively, were synthesised and tested for their pore-forming and transfection capabilities in HEK 293 and undifferentiated mouse ES cells using patch-clamp recording, Ca2+ imaging and flow cytometry. Polymerisation reactions were enhanced by microwaves, and the product sizes were controlled by altering the irradiation time. This method can also be applied to obtain polymers with variable linking chains as shown by the preparation of poly-(1,3-octylpyridinium) salt of 11.9-kDa size. Molecular weights of the final products were determined using ESIMS analysis, which also indicated the products to be amongst the largest macro-cycles ever recorded, up to a 900-membered ring. Anti-bacterial, haemolytic and anti-acetylcholinesterase activities were also reported for the two dodecyl pyridinium polymers. These biological activities are characteristic to the structurally related marine toxin, poly-APS

Impact of pollution on rivers in Montenegro: Ecotoxicological perspective

Montenegrin surface water and groundwater are important for the Balkan Peninsula since they are connected by the transboundary Dinaric Karst Aquifer System with the waters of additional five countries. The pollution from the surface water can rapidly infiltrate in aquifer and endanger this sensible ecosystem and the health of humans through drinking water supply. This chapter gives insights in the pressures of pollution on Montenegrin waters and in a limited literature data regarding freshwater ecotoxicological studies in Montenegro. Also, this chapter provides new ecotoxicological data obtained during survey in 2019, with a focus on the sites which are identified as hotspots of fecal pollution. The highest responses of biomarkers which indicate embryotoxic, genotoxic, and phytotoxic effects in zebrafish embryo test and in roots of Allium cepa were obtained at Ćehotina – downstream of Pljevlja. Similar results were detected at the site downstream Mojkovac at Tara, yet this site is affected by different type of pollution. Genotoxic endpoints in zebrafish stressed out sites on Morača and Lim rivers which are under pressures of fecal pollution. The data in this chapter provides an insight into current status obtained by the ex situ bioassays and indicates need for more comprehensive in situ assessment

DNAqua-Net: Developing new genetic tools for bioassessment and monitoring of aquatic ecosystems in Europe

publisher: Pensoft Publishers articletitle: DNAqua-Net: Developing new genetic tools for bioassessment and monitoring of aquatic ecosystems in Europe journaltitle: Research Ideas and Outcomes copyright: http://creativecommons.org/licenses/by/4.0/ received: 2016-11-27 published: 2016-11-29publisher: Pensoft Publishers articletitle: DNAqua-Net: Developing new genetic tools for bioassessment and monitoring of aquatic ecosystems in Europe journaltitle: Research Ideas and Outcomes copyright: http://creativecommons.org/licenses/by/4.0/ received: 2016-11-27 published: 2016-11-29publisher: Pensoft Publishers articletitle: DNAqua-Net: Developing new genetic tools for bioassessment and monitoring of aquatic ecosystems in Europe journaltitle: Research Ideas and Outcomes copyright: http://creativecommons.org/licenses/by/4.0/ received: 2016-11-27 published: 2016-11-29publisher: Pensoft Publishers articletitle: DNAqua-Net: Developing new genetic tools for bioassessment and monitoring of aquatic ecosystems in Europe journaltitle: Research Ideas and Outcomes copyright: http://creativecommons.org/licenses/by/4.0/ received: 2016-11-27 published: 2016-11-2